1. Field of the Invention
The present invention relates in general to the cellular biological field and, in particular, to a cell cultivating flask and method for using the cell cultivating flask to grow cells.
2. Description of Related Art
Manufacturers of cell cultivating flasks have been for some time trying to design a cell cultivating flask that is more simple and convenient to use than the traditional cell cultivating devices. A cell cultivating flask is basically a device which contains one or more trays that are designed to receive a cell cultivating media and to enable the growth of cells within the cell cultivating media. Examples of two traditional cell cultivating devices are briefly discussed below and described in U.S. Pat. Nos. 5,310,676 and 4,172,013 both of which are hereby incorporated by reference herein.
Referring to FIGS. 1A and 1B (PRIOR ART), there are illustrated two perspective views of a traditional cell cultivating device described in the aforementioned U.S. Pat. No. 5,310,676. The cell cultivating device shown in FIGS. 1A and 1B comprises an outer container or flask 10, which is made from a transparent or non-transparent material, such as glass or plastics material, or from metal. The container 10 is provided with a neck 11 defining a filling opening 12. The neck 11 is formed with outer screw threads 13 for cooperating with inner screw threads of a screw cap 14 by means of which the filling opening 12 may be closed. The outer container 10 has a flat bottom wall 18, a top wall 19, opposite side walls 20, a flat end wall 21, and an opposite end wall 22 on which the neck 11 is formed.
A number of partition wall members 15 having a bottom wall or cell attachment plate 16 and a surrounding peripheral side wall 17 extending upwardly therefrom, is arranged in superposed relationship within the container 10, so as to define cultivating chambers therein. The lower partition wall member 15 arranged adjacent to the bottom wall 18 is supported by feet or spacer members 23 so that a cultivating chamber is defined between the inner surface of the container bottom wall 18 and the lower surface of the bottom wall 16 of the lower partition wall member 15. The spacer members 23 may be formed as integral parts of the walls of the outer container 10 or of the lower partition wall member. An upper cultivating chamber is defined between the inner surface of the container top wall 19 and the upper surface of the adjacent bottom wall 16 of the upper partition wall member 15. Furthermore, a cultivating chamber is defined between each pair of adjacent partition wall members 15. Thus, the cell cultivating device shown in FIGS. 1A and 1B which comprises two partition wall members 15 defines three cultivating chambers therein.
The container end wall 22 has a convex contour so that a manifold chamber 24 is defined between the container end wall 22 and the adjacent side walls 17 of the partition wall members 15. Cut-outs in the peripheral side walls 17 of the partition wall members 15 define overflow openings 26 communicating with the manifold chamber 24.
The outer container 10 may be made from at least two separate container parts, which may, for example, be molded from transparent plastics material. When the partition wall members 15 have been arranged within one of the container parts, such container parts may be sealingly interconnected, for example by interconnecting flanged rim portions of the container parts so as to form a heat seal 27 in a plane being substantially parallel with the bottom walls 16 of the partition wall members 15.
As indicated in broken lines in FIGS. 1A and 1B each of the partition wall members 15 further comprises an equalizing opening 30 formed in its peripheral side wall 17 adjacent to the container end wall 21. The equalizing opening 30 may be a cut-out formed in the rim portion of a curved part of the peripheral wall 17 of each partition wall member. The curved parts of the superposed partition wall members 15 define a transverse tunnel or passage 31 interconnecting the superposed cell cultivating chambers. The tunnel or passage 31 may have any suitable cross-sectional shape, which may, for example, be a semi-circle as shown.
When the cell cultivating device shown in FIGS. 1A and 1B is to be used it is positioned in its upright position shown in FIG. 1A, and the screw closure cap 14 is removed. A suitable amount of cell cultivating medium and cells to be cultivated are poured into the manifold chamber 24 of the container 10 through the filling opening 12 defined by the neck 11. From the manifold chamber 24 of the liquid cell cultivating medium flows into the various cultivating chambers via the overflow opening 26 and through the opening defined between the inner surface of the container bottom wall 18 and the bottom wall 16 of the adjacent partition wall member 15. Now, the level of the cell cultivating media within the cultivating chambers will be the same after a short period of time because the cultivating chambers are all interconnected by the tunnel 31 and the openings 30 formed therein. Provided that the dimensions of the cultivating chambers are substantially the same, these chambers will now contain substantially the same amount of cell cultivating media.
After the filling opening 12 has been closed by the screw cap 14, the flask 10 may then by a quick movement be tilted from the upright position shown in FIG. 1A to the position shown in FIG. 1B in which the container bottom wall 18 is supported in a substantially horizontal position. As the equalizing openings 30 are relatively small, this simple procedure renders it possible to have substantially the same amount of cell cultivating medium placed in all of the now horizontally extending cultivating chambers. This means that a layer of cell cultivating medium containing cells to be cultivated is supported by the container bottom wall 18 and by the bottom walls or cell attachment plates 16 of each of the partition wall members 15. The surfaces of these walls have preferably been subject to a surface treatment allowing good cell attachment.
After expiration of the cultivating period the cell cultivating medium may be poured out through the inlet opening 12 of the neck 11. Thereafter, the cells attached to the bottom walls 16 and 18 may be scraped or flushed out through the inlet or filling opening 12. Alternatively, the container bottom wall 18 or top wall 19, or any of the container side or end walls 20 and 21, respectively, may be cut away or otherwise removed so that the tray members may be taken out from the container 10, whereafter the cultivated cells may be scraped or flushed from the bottom walls 16 and 18.
A main drawback of this traditional cell cultivating device is that a user does not have direct access to each of the cultivating trays. In other words, the user can not fit a pipette of any size onto any cultivating tray to add or remove the cell cultivating media. Of course, it would be desirable to enable the user to have direct access to each of the cultivating trays. Another drawback of this traditional cell cultivating device is that the cell cultivating media often wicks via capillary action between the inner walls of the outer container 10 and the outer walls of the wall members 15. Cells that are trapped in this manner can easily die and contaminate the other cells.
Referring to FIGS. 2A and 2B (PRIOR ART), there are illustrated two sectional front views of a traditional cell culture system described in the aforementioned U.S. Pat. No. 4,172,013. The traditional cell culture system can be used to enable the mass growth of cell by introducing a nutrient medium, cell suspension material etc. through a central feed line 1 serving as supply channel into the lower part of a system of communicating chambers 2. As shown in FIG. 2A, the system is in a vertical position while the nutrient medium is introduced into the communicating chambers 2 which are formed by a number of parallel flat troughs 3. The nutrient medium effectively distributes itself within the communicating chambers 2 with the aid of communicating tubes (not shown) and an central aeration and evacuation channel 4 that can be used to equalize the air pressure within the communicating chambers 2. Thereafter, the nutrient medium, etc., is effectively distributed over the useful surface of the troughs 3 by moving the system into a horizontal position (see FIG. 2B) and by operating valves 5 and 6. In the horizontal position, the cells can be grown on the troughs 3 by controlling the atmosphere within the communicating chambers 2. After completion of the cell growth, the system can be restored to the vertical position which enables the grown cells and nutrient media to flow out through the central supply channel 1.
Like the traditional cell cultivating flask of FIGS. 1A and 1B, the main drawback of this traditional cell culture system is that a user does not have direct access to each of the cultivating troughs 3. In other words, the user can not fit a pipette of any size onto any cultivating trough 3 to add or remove the nutrient medium. Again, it would be desirable to enable the user to have direct access to each of the cultivating troughs 3. Another drawback of this traditional cell culture system is that the troughs 3 are so large that the system must be placed on a xe2x80x9cveryxe2x80x9d flat surface in an incubator or the nutrient media might not sufficiently cover the trough 3 or the nutrient media may be unequal in depth. Moreover, the process of filing the system with the nutrient media, etc., and the process of changing the position of the system from vertical to horizontal and back to vertical are complicated steps that are difficult to perform and if they are not performed correctly the whole cell batch could be contaminated. Accordingly, there is a need for a cell cultivating flask that addresses the aforementioned drawbacks of the traditional cell cultivating devices. In particular, there is a need for a cell cultivating flask that enables a user to have direct access to each cultivating tray therein so as to enable the user to add or remove a cell cultivating media to or from each cultivating tray. These needs and other needs are satisfied by the cell cultivating flask and method of the present invention.
The present invention includes a cell cultivating flask and a method for growing cells within the cell cultivating flask which is configured such that a user can directly access the individual trays or cell growing areas within the cell cultivating flask. Basically, the cell cultivating flask includes a cover located on top of an intermediate tray which is located on top of a bottom tray. The intermediate tray and bottom tray each have a bottom plate and side walls that define a cell growth area. And, the cover and intermediate tray each have a neck with an opening formed therein which enables a user to directly add or remove a cell cultivating media to or from each of the cell growth areas in the intermediate tray and the bottom tray. The cell cultivating flask may have more than one intermediate tray.